In the present study, we developed a split-pot bioassay to screen the indole acetic acid producing rhizobacteria to improve the adventitious root initiation and plant growth promotion in tomato. Based on the ability to produce IAA, 12 isolates were selected among which IRB1 and IRB2 were negative and other rhizobacteria showed a varied level of IAA production (0–84.7 μg/ml) when grown in media supplemented with increasing concentration of L-tryptophan (0–500 μg/ml). Results of split-pot bioassay revealed the capability of IAA producing bacteria to stimulate the growth of adventitious roots on tomato stem which was significantly (P ≤ 0.05) higher than control and IAA negative bacterial treatments. Further enhanced growth of adventitious roots was in accordance with their IAA producing ability. The probability of obtaining consistent results with split-pot bioassay was analyzed by repeating the experiments ten times and subjecting the data to statistical analysis. The results obtained indicated that split-pot bioassay has significantly lesser variations between the individual experiments in comparison with regular bioassay. Further, randomly selected 31 rhizobacteria with unknown identity and characters from the rhizosphere of tomato plants were screened by employing the split-pot bioassay. Isolates UKR8, UKR19, UKR23 and UKR30 were found significant (P ≤ 0.05) in increasing the number and length of adventitious roots and increase in root length, shoot length and fresh weight under greenhouse conditions when compared to regular bioassay. It was also recorded that these bacteria were found to produce IAA to various extent. A possible application of split-pot bioassay in screening IAA producing rhizobacteria and their further application in improving tomato plant growth is discussed in the present study.